Research Summary Nature readily creates and utilizes chemical diversity for the evolution of potent natural products, enzymes with new functions, and even complex systems. Rather than compete with Nature, my laboratory looks to co-opt biological systems to synthesize and evolve chemical diversity by bringing together modern methods in chemical synthesis and DNA technology. The last century saw a revolution in our understanding of the reactivity of small molecules and ability to synthesize small molecules of defined molecular structure, realized as the modern drug industry. My research aims to bring this level of control and understanding to complex biological systems. Manipulation of these biological systems should not only allow us to make new and useful materials on a whole new scale, but also provide fundamental insight into the mechanism of these complex biological systems. Our long-term goal is to understand protein function at the molecular level, looking at isolated proteins in solution, large protein complexes, and finally protein function in biological networks in living cells. Chemical Complementation. Advances in computation and directed evolution hold promise for being able to understand and design protein catalysts with tailor-made structures and functions. Such proteins could be used as materials, reagents, and even therapeutics. The question of how a protein’s primary amino acid sequence dictates its three dimensional fold and function is not resolved and is of fundamental importance to our understanding of living systems and the design and synthesis of higher-order structures. Directed evolution attempts to recapitulate the natural evolution of proteins with new structures and functions, but on an experimentally accessible timescale. Genetic methods have the advantage of DNA encoding, but are limited to the repertoire of chemistry used by nature. Here we have sought to combine the advantages of genetic assays with the flexibility of synthetic chemistry by linking enzyme catalysis to traditional genetic assays for reporter gene transcription via small molecules. The genetics allows us to use DNA encoding, and the small molecule chemistry allows us to readily extend this assay to new chemical reactions. Currently, we are using directed evolution both to ask fundamental questions about the molecular basis for enzyme catalysis and to engineer enzymes with new and useful properties.

Ribosome Chemistry. The ribosomal biosynthetic machinery, a large complex of protein and RNA, is among Nature’s most sophisticated biosynthetic machineries. The ribosome in essence allows template-encoded synthesis of polymers of defined length and composition. Unlike in most biosynthetic machines, substrate recognition is separate from the catalytic center in the ribosome, suggesting it may be particular tolerant to substrate manipulation. Here, our goal is to extend efforts to use synthetic aminoacyl-tRNAs to expand the genetic code, instead to read-out the 64 natural codons with artificial substrates using a purified translation system. This project is in collaboration with Prof. Steve Blacklow and Dr. Tony Forster at Harvard Medical School. Currently, we are using this system for ribosome display of peptidomimetics and to test the adaptor hypothesis, one of the fundamental tenets of translation.

In Vivo Imaging. Finally, in collaboration with Prof. Mike Sheetz in Columbia's Biological Sciences Department, we are developing methods for selectively labeling proteins with small molecules inside the cell. The short-term goal of this project is to provide chemical surrogates to GFP for multi-color tagging and FRET applications. The long-term goal is to extend the power of synthetic chemistry to living systems.